Proteomic tools to study phosphorylation of intrinsically disordered proteins

Area of Science:

• Biochemistry
• Proteomics
• Structural Biology

Background:

• Intrinsically disordered proteins (IDPs) lack defined structures, acting as hubs in cellular networks and driving organelle formation.
• IDPs are prone to post-translational modifications (PTMs), such as phosphorylation, which modulate their function.
• Dysregulation of IDPs and their PTMs is implicated in various diseases.

Purpose of the Study:

• To review analytical approaches for studying IDP phosphorylation.
• To highlight mass spectrometry-based techniques for investigating phosphorylation-dependent conformational changes in IDPs.
• To discuss the potential of advanced mass spectrometry for IDP research.

Main Methods:

• IDP enrichment strategies including strong acid extractions and heat-based pre-fractionation.
• Phosphopeptide and phosphoprotein enrichment and mapping techniques.
• Mass spectrometry-based methods: limited proteolysis, hydrogen-deuterium exchange (HDX), chemical cross-linking, covalent labeling, and ion mobility.

Main Results:

• Discussion of various methods for enriching and analyzing phosphorylated IDPs.
• Overview of techniques to probe IDP conformational dynamics influenced by phosphorylation.
• Emphasis on the utility of ion mobility and electron transfer dissociation in mass spectrometry for IDP studies.

Conclusions:

• Advanced mass spectrometry techniques are essential for studying IDP phosphorylation and conformational alterations.
• Exploiting IDP disorder aids purification and synthetic production for structural studies.
• Technological advancements in mass spectrometry, particularly ion mobility, will enhance insights into IDP biology and disease relevance.